Abstract:
The transport of 35S(-II) by 3 different strains of wine yeasts, Saccharomyces cerevisiae strains R92 and R104 and Saccharomyces chevalieri strain R93 has been investigated.
Chapter one is divided into 4 main sections:
- a general description of the yeast and a discussion of the membrane structure;
- the transport systems in a living membrane;
- a brief literature survey on the transport systems in unicellular organisms;
- the biochemistry of sulphur and the chemistry of sulphide.
The experimental technique used is described in chapter two while chapter three discusses the various tests performed on the experimental technique to check for its validity. Tests performed on the yeasts, to check for their viability, under various experimental conditions are also discussed in chapter three.
The results obtained from the present investigation are given in chapter four and these results are discussed in chapter five.
The uptake of S(-II) has been investigated over a range of pH values (3.1 to 7.8). A pH effect was observed but this effect could neither be related to changes in the H2S (or HS-) concentration with pH nor to the hydrogen ion concentration. Two pH values (3.1 and 7.2) were therefore chosen for the remaining uptake studies.
The transport process of S(-II) appeared to be due to simple diffusion of H2S(aq) and carrier mediated transport of HS-. The kinetic constants Km and Vmax were calculated for the carrier component of the mechanism at pH 7.2 and the permeability coefficient P was calculated for the diffusion of H2S(aq) at pH 3.1 and 7.2. By using these parameters, it was possible to calculate a theoretical initial rate of uptake over a range of extracellular S(-II) concentrations (0 to 50 mmol l-1) at pH 3.1 and pH 7.2. The experimentally determined initial rates were found to agree, within the experimental error, with the theoretical values.
The uptake of S(-II) was not dependent on carbonyl cyanide -m- chlorophenylhydrazone, azide or cyanide ions at pH 3.1, while addition of these species inhibited the uptake of S(-II) at pH 7.2.
Preincubation in glucose, (an energy providing substrate) increased the uptake of S(-II) from a 5 mmol l-1 solution at pH 7.2.
The activation energy (Ea) for the uptake of S(-II) from a 5 mmol l-1 solution at pH 7.2 was higher than at pH 3.1, but when a lower (S-II) concentration was used this difference was not observed. Variation of Ea with extracellular S(-II) concentration was also observed.
Several intermediates of the sulphur reduction pathway were identified by autoradiography following the uptake of S(-II). Their presence was taken as evidence for the influx of 35S(-II).
10% of the 35S was effluxed and this percentage was found to be independent of the efflux media.
The initial rate of uptake of S(-II) and the values of Km for yeast strain R104 (a low sulphide producer) were found to be less than for both strain R92 (a normal sulphide producer) and for strain R93 (a high sulphide producer).